Comparative genomics is a powerful tool for uncovering patterns of mutations within genes targeted for issues in human health. Our research approach utilizes molecular phylogenetic methods and the well-defined evolutionary relationships (based on multiple genetic markers) that unite the 37 species within the cat family Felidae as a reference phylogeny. Using the cat family Felidae as model taxa, we demonstrate different patterns of molecular evolution between genetic markers located on the X- and Y-chromosomes. We are investigating patterns of mutations within four genes associated with varying extent to reproductive fitness. SRY (sex determining region on the Y) is a single copy gene exclusive to the Y chromosome and is the trigger for male sex determination during embryonic development. Phylogenetic analyses indicate that purifying selection may be occurring within SRY and that slower rates of evolution distinguish SRY from its genomic noncoding flanking regions. The other three genes under investigation are single copy X-Y homologues which are fully functional and do not undergo X inactivation in females nor dosage compensation in males. Like SRY, these genes are located within the non-recombining region of the Y (NRY) chromosome, and therefore, do not undergo conventional recombination with their X chromosome counterpart. Originally the NRY was thought to be a "functional wasteland" and these genes were expected to either become specialized for male function, or gradually degenerate from the accumulation of deleterious mutations. By using a phylogenetic approach, we are testing patterns of evolution and change within the NRY genes. Analyses of sequences from intron regions of ZFY (zinc finger on the Y), SMCY (select mouse cDNA on the Y) and UBE1Y (ubiquitin activating enzyme -1 on the Y) within 36 species of Felidae and an additional 22 species sampled from across the order Carnivora reveal the following: 1) faster overall rates of substitution in the Y-chromosome than X-chromosome; 2) an alternate form of recombination, ectopic gene conversion, that may be a mechanism for maintaining function in Y-chromosome genes; and 3) very high phylogenetic signal in defining patterns of speciation. Using the recently published human Y sequence as a "guidepost" we are currently characterizing gene content and order within the domestic cat Y. Using a genomic Y library of domestic cat, we are screening and sequencing single copy Y genes for comparative analyses with the human Y chromosome. These ongoing investigations are integral to understanding the utility and power of sex-linked markers in the assessment of the mode and tempo of speciation; depiction of historical and ongoing patterns of migration; determination of current population structure and viability; and ultimately, characterize those mutational processes affecting genes associated with male reproductive fertility.